ABSTRACT The overall goal of the proposed research is to develop and evaluate new peptide nucleic acid-peptide (PNA- peptide) conjugates for in vivo imaging of the B-cell lymphoma/leukemia-2 (bcl-2) proto-oncogene in non- Hodgkin's lymphoma (NHL). The hypotheses to be addressed in this proposal are 1) that overexpression of bcl-2 mRNA can be detected in vivo in bcl-2 positive, as opposed to negative, NHL tumors, using radiolabeled antisense PNAs conjugated to a peptide for tumor-associated receptor-mediated delivery, and 2) that in vivo imaging of bcl-2 overexpression can predict relapse following conventional chemotherapy in canine lymphoma patients. In aggressive lymphoma, large cohort studies have shown that overexpression of the bcl-2 gene correlates strongly with resistance to radiation and chemotherapy, increased survival of cancer cells, high relapse rate, and poor disease-free and overall survival. A major objective of this project is to determine whether molecular imaging of bcl-2 status in NHL has prognostic significance for relapsed and chemorefractory patients. We will accomplish this goal by three specific objectives. First, evaluating 111In-labeled PNA-peptide conjugates for mRNA targeting in matched cell and animal models of cancer with or without introduction of a the bcl-2 gene, Ramos (bcl-2-negative) and Ramos/bcl-2 (bcl-2-positive) (Specific Objective 1). We will also evaluate an 111In-labeled anti-bcl-2 PNA-peptide conjugate in a veterinary clinical imaging trial, by performing molecular scintigraphy of canine NHL patients to determine whether non-invasive bcl-2 detection can predict relapse (Specific Objective 2). Thirdly, the toxicity of the bcl-2 antisense PNA-peptide conjugate will be evaluated by performing single dose acute toxicity studies of the compound in non-tumor-bearing mice, with the future goal of initiating human imaging studies (Specific Objective 3). A major clinical goal of this proposal is to conduct a prospective imaging study using the radiolabeled anti-bcl-2 PNA-peptide conjugate in canine lymphoma patients during and after remission following conventional chemotherapy, in order to determine whether positive imaging results predict disease relapse. This goal will facilitate translational research in human subjects. Potential Impact on Veterans Healthcare - NHL is among the most common hematopoietic malignancies. The incidence of NHL is increasing, and it is one of three cancers for which mortality has increased substantially during the past decade. NHL represents 4-5% of all new cancer diagnoses and is the fifth leading cause of cancer-related death in America, and approximately 11.5% of its victims are US war veterans. Response of NHL patients to conventional combination chemotherapy is initially good, with 30-35% achieving long-term survival at 10-15 years follow-up. Unfortunately, in most cases, relapse following chemotherapy is inevitable, with transformation to a more aggressive histologic type and development of drug resistance. High bcl-2 expression was more frequently associated with stage III-IV disease and correlated strongly with high relapse rate, reduced disease-free survival, and poor overall survival. Thus, bcl-2 overexpression might be considered a new independent prognostic parameter in NHL, aiding in the identification of patient risk groups. Clinical understanding of the role of bcl-2 in disease progression and treatment response or failure will become increasingly important as drugs that act through mechanisms targeting this proto-oncogene are discovered. Bcl-2-positive patients might respond better to alternative treatments, such as targeted immunotherapy, radioimmunotherapy (RIT), or antisense therapy, all of which act through mechanisms that down-regulate bcl- 2. This goal is central to the Veterans Health Administration (VHA) mission of providing primary and specialized care to cancer patients who are veterans. PUBLIC HEALTH RELEVANCE: The proposed research focuses on non-Hodgkin's lymphoma (NHL) as one of the most common hematopoietic malignancies. NHL is the fifth leading cause of cancer-related death in America, and approximately 11.5% of its victims are U.S. war veterans. The goal of the proposed research is central to the VHA mission of providing primary and specialized care to cancer patients who are veterans.